The long-term goal of this proposal is to understand how neurotransmitters act through G protein-coupled receptors to modulate the activities of neurons. Our studies focus on a neurotransmitter signaling pathway that regulates activities of the neurons and muscles of the egg-laying system of C. elegans. We have identified mutations that disrupt the regulation of egg-laying behavior and used them to identify and study genes encoding components of this G protein signaling pathway. The genes analyzed so far have close homologs expressed in the mammalian brain, suggesting that C. elegans will prove a useful model for understanding neurotransmission through G proteins in humans. The first major aim of this proposal is to systematically exploit the molecular genetic system we have developed to analyze this neurotransmitter signaling pathway. We will carry out saturating genetic screens to identify as many of the signaling genes as possible. We will clone and molecularly analyze these genes. Our goal is to eventually reduce our understanding of this G protein signaling pathway to a biochemical level. The potential of this approach is illustrated by the fact that we have already used it to identify a protein, EGL-10, that inhibits signaling by a heterotrimeric G protein in the C. elegans egg-laying system. EGL-10 is a prototype for a large family of proteins we identified in worms and humans that we have named the "regulator of G protein signaling" (RGS) proteins. These RGS proteins may regulate may or all G protein signaling pathways. The second major aim of this proposal is to determine why there are so many RGS proteins. Are they simply redundant, or does each have a specific biological function, perhaps because each is restricted to interacting with a particular G protein or set of G proteins? We will examine this issue by determining the genetic functions and expression patterns of the 11 worm RGS proteins and by studying the biochemical specificity of the interactions between C. elegans RGS and G proteins in vitro. this analysis should reveal the logic by which the family of RGS proteins regulates multiple signaling pathways in the worm. Because many C. elegans G proteins and RGS proteins are closely related to corresponding human homologs, our analysis should shed light on analogous human signaling pathways and direct their further study.